The biogenesis of the chloroplast has been described in great detail. However, the transcriptional activities of this DNA have not been described, and very little is known about transcriptional regulation of organelle gene expression. Our previous research of chloroplast gene expression has indicated that this genome is expressed at different rates during different stages of cell growth and development (Matsuda and Surzycki, 1979). We have also established that five proteins for the 30S chloroplast ribosomal subunit are encoded in chloroplast DNA (Surzycki and Chandler, 1979). In general, our work has indicated that all transcription units in the chloroplast genome can be classifield into three categories: (a) transcription units expressed predominantly at some but not all stages, (b) transcription units expressed continously and, (c) transcription units not expressed during cell growth and greening. The proposed research will be a continuation of our previous studies and will concentrate on the following problems: (1) We will construct a detailed map of chloroplast DNA transcription on the level of individual transcription units. Cloned fragments of chloroplast DNA, EM techniques (e.g. R-loop) and Sl nuclease mapping will be used. We hope to determine the structure of transcription units, the level of their expression at various stages of cell development and the amount of RNA from each unit that accumulates in the cell. We will also determine if split genes are present in the chloroplast genome. (2) We will extend the analysis of transcription to the processes of gamete formation and zygote maturation. (3) We will study the number and the location of the genes in chloroplast DNA which produce RNA containing poly(A) sequences. (4) We will locate the exact positions of the five chloroplast ribosomal protein genes on the physical map of DNA. The method of hybridization-arrested translation using cloned fragments of chloroplast DNA will be used. This analysis will be extended to the ribosomal proteins of 50S subunit. (5) We will clone the appropriate chloroplast DNA fragments and establish a cloned library of this genome. This will facilite the determination of which genes are encoded in chloroplast DNA and how their expression is regulated.